A large ribonucleoprotein complex called the ribosome is responsible for several steps of protein synthesis in all organisms. In bacteria, regulation of translation begins at initiation. Despite the availability of structural information, we still lack a clear understanding of how the ribosome encounters folded mRNA structures during translation initiation.
A general mechanism for initiation could involve a ribosome standby binding to single-stranded regions of the mRNA in the vicinity of the ribosome binding site, representing the first docking step and thus providing a general mechanism for initiation. This standby model describes the paradox of high translation rates from highly structured mRNAs with a sequestered RBS. Until now, little attention has been paid to standby complexes, particularly those of structured mRNAs.
Riboswitches are structured cis-acting RNA elements located in the 5’-untranslated region of mRNA that regulate gene-expression at the level of transcription, RNA cleavage and translation in response to binding of a cognate ligand.
The direct regulation by riboswitches operates by interfering with the formation of translation initiation complexes.
However, a structural description of the interaction between riboswitches and ribosomes during translation initiation is currently missing. It therefore has remained elusive how further structured elements can be fully accommodated within the initiation complex. We investigate the influence of the 30S ribosome on the function of the adenine-sensing riboswitch from Vibrio vulnificus by NMR spectroscopy. Surprisingly, the adenine-induced allosteric switch leading to an opening of the RBS is insufficient for efficient translation initiation. Additional stable structured elements around the initiation region prevent mRNA accommodation in the ribosome decoding channel.
Our results show that the full activity of the riboswitch is only obtained upon concerted interaction with adenine and ribosomal proteins. The RNA chaperone activity of the ribosomal protein S1 is needed for melting of secondary structures that would otherwise preclude complex formation.